The amide transfer reaction catalyzed by amide transferases is poorly understood in spite of its importance in various biosynthetic pathways. Three amide transferases, namely, FGAR amidotransferase, XMP aminase, and NAD synthetase will be investigated. Preliminary studies have shown that FGAR amidotransferase (from chicken liver) catalyzes the FGAR, ATP, Mg 2 plus dependent hydrolysis of gamma-glutamyl hydroxamate, gamma-glutamyl hydrazide and gamma- glutamyl methoxyamide in addition to the amide bond of glutamine. The released substituted-amide group may be transferred to the keto function of FGAR to form FGAM derivatives. Further studies on the nature of gamma-glutamyl substituted-amides as alternate nitrogen donors are being carried out. Preliminary studies on the active site of FGAR amidotransferase indicate that iodoacetate, N-ethylmaleimide (NEM), D- and L-alpha-bromopropionate can selectively alkylate the glutamine-site of the enzyme. Isolation and determination of the sequence of the peptide containing C14-iodoacetate labeled glutamine-site are in progress. Upon modification of the enzyme by NEM, a biphasic inactivation of the enzyme was observed. The enzyme is inactivatted at different rates by the antipodes of alpha-bromopropionic acid. The L- (plus) antipodes react faster. The nature of NEM modification and the relationship between the stereoselective chemical reactivity and stereospecificity of the enzyme will be investigated. Procedures will be developed for the purification of XMP aminase and NAD synthetase to a homogeneous state from suitable sources. The pure enzymes will be used as tools for the investigation of amide transfer reaction. Furthermore, the role of XMP aminase and NAD synthetase in the control of the biosynthesis of GMP and NAD, respectively, will be studied.